The long-term objective of this research proposal is to understand the relationship between Cdc7p and the Mcm2-7 complex in DNA replication. Presently, it is not well understood how the eukaryotic Mcm complex is regulated, and exactly what role Cdc7 plays in this regulation. Our laboratory has isolated a recessive mutation in S. cerevisiae Mcm5p, called mcm5-bob1, which allows for viability in the absence of the normally essential Cdc7/Dbf4 complex. The hypothesis is that mcm5-bob1 mutation adopts an altered conformation (in the absence of Cdc7p), which may mimic a phosphorylation event normally executed by Cdc7p. This proposal will help elucidate the relationship and regulation of the Mcm2-7 complex by Cdc7p. The specific aims of this proposal are to: 1) To identify new gene products which interact with Cdc7 and the Mcm complex in DNA replication, using random transposon mutagenesis, 2) To perform a structure/function analysis of the yeast Mcm complex, using the M. thermoautotrophicum Mcm protein as a structural model, and 3) To analyze the molecular functions of mutant S. cerevisiae Mcm complexes using point mutations in yeast Mcm5 and to test the stability of the mutant Mcm complexes using co-IP and 2-hybrid analysis. [unreadable] [unreadable]